The long-term success of cardiac transplantation is currently limited by the high incidence of transplant-related coronary artery disease (TCAD) (Hosenpud et al., 1996, J. Heart and Lung Transplantation 15:655). This complication may be related to the recipient's ongoing immune response against donor major histocompatibility complex (MHC) antigens since long-term allograft survival correlates directly with the number of donor-recipient human leukocyte antigen (HLA) matches (Kormos, et al., 1992, J. Heart and Lung Transplantation 11(3:S104–110; Kerman, et al., 1994, Transplantation 1994, 57(6): 884–8); Smith, et al., 1995, Lancet 346: 1318–22; Constanzo-Nordin, M. R., 1992, J. Heart and Lung Transplantation 11: S90–103) and inversely with the development of circulating antibodies against donor HLA molecules (Suciu-Foca, et al., 1991, Transplantation 51:716–724; Hess, et al., 1983, Circulation 68:94–101; Rose, et al., 1992, J. Heart and Lung Transplantation S120–123). Moreover, since donor/recipient HLA-DR mismatching is associated with increased cardiac allograft rejection episodes (DeMattos, et al., 1994, Transplantation 57(4)626–630; Kirklin, et al., 1994, Transplantation 13(4): 583–95; Keogh, et al., 1995, J. Heart and Lung Transplantation 14(3):444–51), TCAD may be the end result of recurrent or persistent allograft rejection (Constanzo-Nording, M. R., 1992, J. Heart and Lung Transplantation 11: S90–103).
In order to identify patients at high risk of having a positive donor-specific cross-match, cardiac transplantation candidates are prospectively tested for anti-HLA antibodies against lymphocytes from a panel of volunteers representative of the major HLA allotypes, collectively referred to as measurements of panel-reactive antibodies (PRA). In addition to predicting an increased likelihood of donor-specific anti-HLA antibodies and a consequent risk of early graft failure related to humoral rejection, several studies have shown that high levels of pretransplant PRA in cardiac allograft recipients are associated with adverse post-transplant outcome when compared to patients with low or negative reactivity (Smith et al., 1993, Transplant Immunol. 1:60–65). High PRA levels have been associated, in some studies, with increased frequency of acute cellular rejections, decreased long-term graft survival, and increased mortality (Lavee et al., 1991, J. Heart and Lung Transplantation 10:921–930; Loh et al., 1994, J. Heart and Lung Transplantation 13:194–201). Moreover, the onset of accelerated coronary artery disease (CAD) in cardiac transplant recipients, the major limitation to long-term graft survival, has been associated with the presence of anti-HLA antibodies (Hess et al., 1983, Circulation 68:94–101; Rose et al., 1989, Surgery 106:203–208; Suciu-Foca et al., 1991, Transplantation 51:716–724). Since accelerated CAD in these patients may be a consequence of cumulative episodes of high-grade cellular rejections, it is possible that this association may actually reflect a relationship between anti-HLA antibodies and acute cellular rejection.
The only consistently reliable method for diagnosis of cardiac allograft rejection in patients who have received a transplant is the endomyocardial biopsy (EMB). The probability of progression from a negative or low-grade EMB to a high-grade biopsy is greatest during the first six months following transplantation (Rizeq, et al., 1994, J. Heart and Lung Transplantation 13(5):862–868; Winters, et al., 1995, Circulation 91:1975–1980; White, et al., 1995, J. Heart and Lung Transplantation 14:1052; Billingham, M., 1990, J. of Heart and Lung Transplantation 9:77). Since the EMB can only diagnose established rejection episodes, and the procedure has several drawbacks, including risk of complications, high cost, sampling error, and potential variation in interpretation, it would be highly desirable to have non-invasive modalities to prospectively identify patients at high risk of progression from a low to high EMB grade. Although various non-immunologic modalities, including measurement of hemodynamic parameters (Valentine, et al., 1991, J. Heart and Lung Transplantation 10:557), radionuclide scanning (Kemkes, et al. 1992, J. Heart and Lung Transplantation 10:557), and magnetic resonance imaging (Baumgartner, 1986, J. Heart and Lung Transplantation 5:419; Wisenberg, 1987, American Journal of Cardiology, 60:130), have shown good correlation with established high-grade rejections, none have demonstrated sufficient predictive value to be included in routine clinical management. Among immunologic assays, measurement of panel-reactive anti-HLA antibodies (George, et al., 1995, J. Heart and Lung Transplantation 14:856–864; Smith, et al., 1992, Transplantation 53:1358–62; Zavazava, et al., 1993, Tissue Antigens 42:20–26), detection of IL-2 activated T cells in the allograft using a 48-hour IL-2 dependent lymphocyte growth assay (LGA) (Fischer, et al., 1995, J. Heart and Lung Transplantation 14:1156–1161), recipient sensitization to donor derived HLA peptides (Liu, et al., 1993, J. Experimental Medicine 177:1643–1650), (Liu, et al., 1996, J. Clinical Investigation 98:1150–1157; Tugulea, et al., 1997, Transplantation 64:842–847; Ciubotariu, 1998, J. Clinical Investigation 101:398–405), and lack of induction of donor-specific hyporeactivity, as measured by recipient T cell proliferation against donor cells (Markus, et al., 1993, Cell Transplantation 2:345–353; Zeevi, et al., 1995, Transplantation 59:616–620; Creemers, et al., 1997, Nephron 75:166–170) have been shown to correlate with episodes of established cellular rejection, however the predictive value of these assays has not been extensively evaluated.
Currently, five year allograft survival is estimated to occur in 70% of cardiac transplantation patients; with approximately 40% of patients experience a high-grade rejection episode in the first year after transplant. For transplants involving other tissues, long-term graft survival continues to be a problem. For example, in kidney and liver the ten year survival rate is approximately 50%; for lung and pancreas the three year survival rate is approximately 50%.
The immunological basis for transplant rejection is the subject of extensive research. Cumulative experimental data in rodent models suggest that initiation of allograft rejection is predominantly a CD4 T cell-dependent process, and that there may not be an absolute requirement for CD8 cells (Krieger, et al., 1996, J. Experimental Medicine 184:2013–2018). Moreover, long-term graft acceptance appears to be associated with reduced direct recognition of donor alloantigens by recipient CD4 T cells (Zeevi, et al., 1995, Transplantation 59:616–620; Creemers, et al., 1997, Nephron 75:166–170). Donor-specific hyporesponsiveness can be augmented by infusion of donor bone marrow cells at the time transplantation (Zeevi, et al., 1995, Transplantation 59:616–620), and may be accompanied by persistent microchimerism (Starzl, et al., 1994, Progress in Liver Diseases 12:191–123), suggesting that recipient CD4 T cells can be rendered tolerant to direct allostimulation by donor leukocytes. However, since donor-type microchimerism has not been found to correlate well with either acute or chronic allograft rejection (Schlitt, et al., 1994, Lancet 343:1469–1471), mechanisms other than direct allorecognition may significantly impact on allograft rejection. Recent evidence has emerged that over time the indirect pathway of CD4 T cell activation plays an increasingly important role in the development of acute and chronic allograft rejection (Liu, et al., 1993, J. Experimental Medicine 177:1643–1650). This may be a consequence of continuous shedding of donor alloantigenic HLA peptides and their processing by host antigen-presenting cells (APCs) such as macrophages and B cells. Primary rejections appear to be invariably accompanied by recipient T cell recognition of a dominant HLA-DR allopeptide presented by self-APCs (Liu, et al., 1996, J. Clinical Investigation 98:1150–1157; Tugulea, et al., 1997, Transplantation 64:842–847), whereas recurrent rejections, as well as the onset of TCAD, appear to be accompanied by inter- and intra-molecular spreading and T cell recognition of multiple donor HLA-DR alloantigenic determinants (Tugulea, et al., 1997, Transplantation 64:842–847; Ciubotariu, 1998, J. Clinical Investigation 101:398–405). This diversification of the immune response has been postulated to be a result of activation of antigen-specifics B cells by soluble MHC class II products, particularly HLA-DR molecules, and the subsequent efficient presentation of multiple HLA-DR allopeptides by self B cells to CD4 T cells (Vanderlugt, et al., 1996, Current Opinion in Immunology 8:831–836; Mamula, et al., 1993, Immunology Today 14:151–154; Reed, et al., 1996, Transplantation 61:556–572).